High CO2 alters the hypoxia response of the Pacific whiteleg shrimp (Litopenaeus vannamei) transcriptome including known and novel hemocyanin isoforms.

Acclimation to low O2 in many organisms involves changes at the level of the transcriptome. Here we used high-throughput RNA sequencing (RNA-Seq) to explore the global transcriptomic response and specific involvement of a suite of hemocyanin (Hc) subunits to low O2 alone and in combination with high CO2, which naturally co-occurs with low O2. Hepatopancreas mRNA of juvenile L. vannamei exposed to air-saturated water, low O2, or low O2/high CO2 for 4 or 24 h was pooled, sequenced (HiSeq 2500) and assembled (Trinity: 52,190 contigs) to create a deep strand-specific reference transcriptome. Annotation of the assembly revealed sequences encoding the previously described small Hc subunit (HcS), and three full-length isoforms of the large subunit (HcL1-3). In addition to this, a previously unidentified full-length Hc subunit was discovered. Phylogenetic analysis demonstrated the subunit to be a ß-type Hc subunit (denoted HcB), making this the first report of a ß-type hemocyanin subunit in the Penaeoidea. RNAs of individual shrimp were sequenced; regulated genes identified from pairwise comparisons demonstrated a distinct pattern of regulation between prolonged low O2 and low O2/high CO2 treatments by GO term enrichment analysis (Roff-Bentzen, P < 0.0001), showcasing the stabilization of energetically costly translational machinery, mobilization of energy stores, and downregulation of the ubiquitin/proteasomal degradation machinery. Exposure to hypoxia for 24 h resulted in an increase in all of the full-length hemocyanin subunits (HcS, HcL1, HcL2, HcL3, and HcB). The addition of CO2 to hypoxia muted the transcriptomic response of all the Hc subunits to low O2, except for the ß-type subunit.

Recovery from hypoxia and hypercapnic hypoxia: impacts on the transcription of key antioxidants in the shrimp Litopenaeus vannamei.

Estuarine waters are prone to regular bouts of low oxygen (hypoxia) and high carbon dioxide (hypercapnia). In vertebrates, tissue hypoxia followed by reoxygenation can generate high levels of reactive oxygen species (ROS) that exceed cellular antioxidant capacity, leading to tissue damage. Here we quantified the expression of several antioxidant genes in the hepatopancreas of Pacific whiteleg shrimp, Litopenaeus vannamei, after exposure to hypoxia or hypercapnic hypoxia for 4h or 24h followed by recovery in air-saturated water (normoxia) for 0, 1, 6 or 24h, as compared to time-matched controls maintained only in normoxia. Transcripts of cytoplasmic Mn-superoxide dismutase (cMnSOD), glutathione peroxidase (GPX) and peptide-methionine (R)-S-oxide reductase (MsrB) increased after 4h exposure to either hypoxia or hypercapnic hypoxia; these elevated transcript levels persisted longer in animals recovering from hypercapnic hypoxia than hypoxia alone. cMnSOD transcripts generally increased, but GPX, MsrB, glutathione-S-transferase (GST), and thioredoxin 1 (TRX-1) decreased or did not change in most long-term (24h) treatment-recovery groups. Thus, the transcriptional responses of several antioxidant genes during recovery from tidally-driven hypoxia and hypercapnic hypoxia decrease or are muted by more persistent exposure to these conditions, leaving L. vannamei potentially vulnerable to ROS damage during recovery.